This goal of this project is to improve the immune function of "humanized" mice in order to provide a better model for testing candidate vaccines for HIV/AIDS. Over the past 20 years, techniques for generating human immune function following transfer of human cells to immunodeficient mice have gradually improved. This project begins with the best of the current models, transfer of human CD34-positive hematopoietic progenitor cells to mice with genetic disruption of Rag-2 and IL-2 receptor common gamma chain genes, and systematically evaluates strategies to improve the function of the many engrafted human B lymphocytes observed in this model. The project specifically focuses on two subsets of human B cells, the B1 subset responsible for "natural antibodies" and the marginal zone (MZ) B2 subset responsible for antibodies to carbohydrate antigens, because we are evaluating glycoconjugate vaccines (produced by Dennis Burton and collaborators) that target carbohydrates on the envelope spike of HIV-1. Recent advances in understanding the differentiation of B1 and MZ B cells provide the rationale for supplying supporting stromal cells and differentiation factors that should enhance the survival and function of these human B cells. The deficits recognized in the current humanized mouse model, absence of human follicular dendritic cells and human T cells that are positively selected on mouse epithelial cells, will be repaired by additional cell transplants. An "artificial lymph node" approach that is more amenable to replacing missing supporting cells will also be used to more rapidly evaluate which cell type is critical for fully functional immune responses. This research proposal is directly responsive to RFA-AI-07-015 in that it attempts to characterize B1 and MZ B cell responses to carbohydrates that shield HIV-1 from attack by conventional antibodies targeting protein components of the envelope. Successful execution of the project specific aims will result in a humanized mouse model capable of rapidly screening multiple HIV-1 candidate vaccines for both immunogenicity and protection against HIV-1 infection. This would speed vaccine development and reduce reliance on primate challenge models. There are many challenges to developing an effective vaccine to prevent HIV/AIDS, and one way to more effectively confront these challenges is to develop a small animal model system that accurately reflects the human immune response to candidate vaccines. After many years of work, that goal seems close. This proposal will attempt to fix the few remaining problems in "humanized mice", and use them to test a new AIDS vaccine decorated with sugar groups from the virus.